Large-scale production and purification of biological compounds

Shiloach, Joseph

Abstract

A significant portion of the Labs activities is the production of high-quality and unique biologicals that are not available from commercial sources. On average, the Unit performs between 150 to 200 processes per year, which include fermentation of bacteria, yeast, insect cells, and mammalian cells (in volumes ranging from 2 to 200 liters), and recovery and purification of biomolecules, such as proteins and polysaccharides, from different sources. The following are examples of processes performed: growth of bacteria like Escherichia coli, (recombinant and native), Salmonella, Shighella, Staphylococcus, Haefmofillus influenzae, Bortedella pertussis, Bacillus Anthracis, and various recombinant yeast strains like Saccharomyces cerevisiae and Pichia pastoris. In addition, mammalian cells such as HeLa, CHO, and HEK 293, and insect cells like Sf 9, and High five for transient expression of recombinant proteins were propagated. Several proteins were produced and purified in gram quantities. Examples are: recombinant Pseudomonas aeruginosa exotoxin A, recombinant human adiponectin and recombinant human alpha 1 proteinase inhibitor. ? The various products were needed for a number of collaborative research projects: human adiponectin was made for biological studies, human alpha 1 proteinase inhibitor for glycosylation studies, and several proteins were produced for structural studies among them Toll proteins and G protein-coupled receptors.

Funding Agency

• Agency: National Institute of Health (NIH)
• Institute: National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
• Type: Intramural Research (Z01)
• Project #: 1Z01DK015500-47
• Application #: 7733958
• Support Year: 47
• Fiscal Year: 2008
• Total Cost: $265,540

Institution

• Name: National Institute of Diabetes and Digestive and Kidney Diseases
• Country: United States

Publications

Chill, Liat; Trinh, Loc; Azadi, Parastoo et al. (2009) Production, purification, and characterization of human alpha1 proteinase inhibitor from Aspergillus niger. Biotechnol Bioeng 102:828-44

Phue, Je-Nie; Kedem, Benjamin; Jaluria, Pratik et al. (2007) Evaluating microarrays using a semiparametric approach: application to the central carbon metabolism of Escherichia coli BL21 and JM109. Genomics 89:300-5

White, Jim F; Grodnitzky, Justin; Louis, John M et al. (2007) Dimerization of the class A G protein-coupled neurotensin receptor NTS1 alters G protein interaction. Proc Natl Acad Sci U S A 104:12199-204

Azurmendi, Hugo F; Vionnet, Justine; Wrightson, Lauren et al. (2007) Extracellular structure of polysialic acid explored by on cell solution NMR. Proc Natl Acad Sci U S A 104:11557-61

Karnaukhova, Elena; Ophir, Yakir; Trinh, Loc et al. (2007) Expression of human alpha1-proteinase inhibitor in Aspergillus niger. Microb Cell Fact 6:34

Jaluria, Pratik; Betenbaugh, Michael; Konstantopoulos, Konstantinos et al. (2007) Application of microarrays to identify and characterize genes involved in attachment dependence in HeLa cells. Metab Eng 9:241-51

Kim, Chul-Hee; Pennisi, Patricia; Zhao, Hong et al. (2006) MKR mice are resistant to the metabolic actions of both insulin and adiponectin: discordance between insulin resistance and adiponectin responsiveness. Am J Physiol Endocrinol Metab 291:E298-305

Bell, Jessica K; Botos, Istvan; Hall, Pamela R et al. (2006) The molecular structure of the TLR3 extracellular domain. J Endotoxin Res 12:375-8

Trinh, Loc; Phue, Je-Nie; Jaluria, Pratik et al. (2006) Screen-less expanded bed column: new approach for the recovery and purification of a malaria transmission blocking vaccine candidate from Pichia pastoris. Biotechnol Lett 28:951-8

Jin, Zhigang; Bohach, Gregory A; Shiloach, Joseph et al. (2005) Conjugates of group A and W135 capsular polysaccharides of neisseria meningitidis bound to recombinant Staphylococcus aureus enterotoxin C1: preparation, physicochemical characterization, and immunological properties in mice. Infect Immun 73:7887-93